Chinese traditional medicine Rhizoma Anemarrhenae is the rhizoma of the Anemarrhena asphodeloides Bge, (Liliaceae), widely distributed in Hebei, Neimenggu, Shanxi province, northeast of China and some other areas. Due to the heat-clearing and fire-purging function and the action of promoting the production of body fluid and nourishing the lung, it was frequently used in Traditional Chinese Medicine clinic. The main components of Anemarrhena asphodeloides Bge. are steroidal saponins, together with flavones, oligosaccharides, polysaccharides and fatty acids, et, al. Pharmacological studies show that it has antibiosis, antiviral, pyretolysis, anti-diabetic, conscious-sedation, inhibition of platelet aggregation, anticancer and radioprotective effects, etc.
Timosaponin BII, also named Prototimosaponin AIII, is the main component of rhizoma of Anemarrhena asphodeloides Bge. It has the structure of (25S)-26-O-β-D-glucopyranosyl-22-hydroxy-5β-furostane-3β, 26-diol-3-O-β-D-glucopyranosyl-(1→2)-β-D-galactopyranoside. It has the following formula:

Timosaponin BII was first isolated by Toshio KAWAZAKI in 1963, but he didn't give its structure. In 1991, Seiji NAGUMO first elucidated the structure of Timosaponin BII (Seiji NAGUMO et al, YAKUGAKU ZASSHI, 1991; 111 (1) 306-310). After that Noboru Nakashima (NOBORU NAKASHIMA et al, Journal of Natural Products, 1993; 56(3):345-350), Bai-ping Ma (Bai-ping Ma et al, Acta. Pharm. Sin. 1996; 31 (4): 271-277), Masayasu Kimula (Masayasu KIMURA et al, Biol. Pharm. Bull, 1996; 19 (7): 926-931) and Jian-ying Zhang (Jian-ying ZHANG et al, Clinica Chimica Acta, 1999; 289: 79-88) reported the isolation and activity of Timosaponin BII. Researches indicated that Timosaponin BII has the activities of antidiabetic, inhibition of platelet aggregation, scavenging of free radical and anti-dementia. In almost all the prior art references, the Timosaponin BII was prepared by n-butanol extraction, normal phase silica gel column chromatography, macroporous resin column chromatography and HPLC. But it is ease to emulsify in n-butanol extraction, the amount of sample is limited on silica gel column chromatography, it is difficult to reproduce silica gel, it is difficult to recover the eluent of chloroform-methanol-water. Therefore, the reported methods usually had a difficult process, low yield, low purity and not amenable to industrialized preparation. Even using macroporous resin and reversed-phase liquid chromatography, the eluent of methanol-water will result in the C-22 hydroxyl methoxylation of Timosaponin BII to form a certain fraction of Timosaponin BI (Seiji NAGUMO et al., YAKUGAKU ZASSHI, 1991; 111(1): 306-310). So these methods would waste some samples and could only generate the mixture of Timosaponin BII and Timosaponin BI, and it is too difficult to get the pure Timosaponin BII.

For these reasons, it is necessary to improve the preparative method of Timosaponin BII.